Studies on crystallization and cross-linking of lipase for biocatalysis

Abstract

The development of robust biocatalysts with increased stability and activity is a major challenge to industry. A major breakthrough in this field was the development of cross-linked enzyme crystals with high specificity and stability. A method is described to produce micro crystals of CLEC lipase, which is thermostable and solvent stable. Lipase from Burkholderia cepacia was crystallized using ammonium sulfate and cross-linked with glutaraldehyde to produce catalytically active enzyme. The maximum yield of CLEC was obtained with 70% ammonium sulfate and cross-linked with 5% (v/v) glutaraldehyde. SEM studies showed small hexagonal-shaped crystals of 2-5 microm size. CLEC lipase had improved thermal and reuse stability. It is versatile, having good activity in both polar and nonpolar organic solvents. CLEC lipase was coated using beta cyclodextrin for improving the storage and reuse stability. CLEC was successfully used for esterification of Ibuprofen and synthesis of ethyl butyrate.

Preparation and characterization of guar gum nanoparticles

“…The dialysis was continued for 24 h at 30 ± 2 • C. Enzyme functionalized nanoparticles (EFNP) were prepared by sequentially mixing hydrolyzed GG (5 ml), Lipase PS (0.5 ml), Triton X-100 (0.1 × 10 −3 mM, 5 l), 50% aqueous polyethyleneimine solution (10 l), IPA (500 l), 0.1% TPP (500 l) or boric acid and finally 25% glutaraldehyde (10 l) as crosslinker. The solution was vortexed and sonicated for 5 min at room temperature to get uniform NPs and the lipase activity was estimated [14]. The average particle size (hydrodynamic diameter, Zaverage) and polydispersity of GG NPs were determined by light scattering experiment (PCS) using 3000 HSA Zetasizer equipped with He-Ne laser ( = 633 nm).…”

Cross-linked enzyme crystals of organophosphate hydrolase for electrochemical detection of organophosphorus compounds

“…The concentration of protein in the soluble crude enzyme and CLEC was determined using Lowry's method according to Abraham et al 2004Rajan and Abraham 2007). Bovine serum albumin (BSA) was used as the standard.…”

Carrier-free immobilized enzymes for biocatalysis

“…Crystal size and retention of specific activity could be tuned in some cases by variation of the conditions for crystallization and cross-linking (Margolin 1996). As shown for Burkholderia cepacia lipase, coating of CLECs with surfactants or b-cyclodextrin could contribute to further enhancement of shelf-life and operational stability (Rajan and Abraham 2008). The scope of the CLEC technology is currently limited by the requirement for a successful crystallisation of the (purified) target enzyme.…”